Faster isolation of recombinant factor VIII SQ, with a superporous agarose matrix

• Published in: Journal of Chromatography A Vol. 840; no. 1; pp. 39 - 50
• Main Authors: Pålsson, Eva, Smeds, Anna-Lisa, Petersson, Anna, Larsson, Per-Olof
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 23-04-1999; Elsevier
• Subjects: Biological and medical sciences; Biotechnology; Chemical Engineering; Chromatography, Affinity - methods; Coagulation factors; Comparison with commercial adsorbent; Culture Media, Conditioned - chemistry; Engineering and Technology; Factor VIII; Factor VIII - isolation & purification; Fluorescein-5-isothiocyanate; Fundamental and applied biological sciences. Psychology; Health. Pharmaceutical industry; Immunoaffinity chromatography; Immunologic Techniques; Industrial applications and implications. Economical aspects; Kemiteknik; Production of active biomolecules; Proteins; Recombinant Proteins - isolation & purification; Sensitivity and Specificity; Sepharose; Staphylococcal Protein A; Teknik; Proteins; Agarose, superporous; Ligand distribution; Immunoaffinity chromatography; Factor VIII; Human; Purification; Recombinant protein; Coagulation factor; Porous material; Agarose; Breakthrough curve
• ISSN: 0021-9673
• DOI: 10.1016/S0021-9673(99)00209-5

A superporous agarose matrix was compared with a corresponding homogenous matrix in the isolation of recombinant factor VIII SQ (r-VIII SQ) by immunoaffinity chromatography. As a reference, the commercially available Sepharose FastFlow, used for a similar purification in the industry, was also evaluated. Breakthrough curves are described for flows between 50 and 400 cm/h with pre-purified r-VIII SQ and with cell culture broth. The superporous gel gave the best performance and a 1000-fold purification was obtained in a one-step procedure. The superporous matrix made it possible to increase the throughput about four-fold, presumably due to its better mass transfer properties. The importance of the ligand distribution profile is discussed based upon immunofluorescence microscopy data.